doi:10.1182/blood-2006-07-034348Prepublished online November 30, 2006;   

 Daniel R CourielGhosh, Joyce Neumann, Yvonne Hsu, Jorge De Jesus, Muzaffar H Qazilbash, Richard E Champlin and Rima M Saliba, Marcos de Lima, Sergio Giralt, Borje Andersson, Issa F Khouri, Chitra Hosing, Shubhra Hyper-acute GVHD: risk factors, outcomes, and clinical implications

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HYPER-ACUTE GVHD: RISK FACTORS, OUTCOMES, AND CLINICAL IMPLICATIONS

Rima M. Saliba, Marcos de Lima, Sergio Giralt, Borje Andersson, Issa F. Khouri, Chitra

Hosing, Shubhra Ghosh, Joyce Neumann, Yvonne Hsu, Jorge De Jesus, Muzaffar H.

Qazilbash, Richard E. Champlin, and Daniel R. Couriel

From the Department of Stem Cell Transplantation and Cellular Therapy, The University of

Texas M. D. Anderson Cancer Center, Houston, TX 77030, U.S.A.

R.M.S. designed and performed data analysis and wrote paper; M.d.L., S.G., R.E.C. helped to write the paper; B.A., I.F.K.

contributed analytical tools; C.H., M.H.Q. provided vital comments; S.G. performed data management; J.N., Y.S. provided

patient care in GVHD clinic; J.D.J. was study coordinator; D.R.C designed and performed research and wrote paper.

Scientific heading: Clinical Observations, Interventions, and Therapeutic Trials

Running title: Incidence and risk factors for hyper-acute GVHD

Key words: acute graft-versus-host disease, allogeneic bone marrow transplantation

Partially presented at the 46th annual meeting of the American Society of Hematology, San Diego, CA, December 7, 2004

Corresponding author:

Daniel R. Couriel, MD

Department of Stem Cell Transplantation and Cellular Therapy,

The University of Texas M. D. Anderson Cancer Center,

1515 Holcombe Blvd, Unit 0423,

Houston, TX 77030

Tel: 713-745-2803

Fax: 713-794-4902

E-mail: dcouriel@mdanderson.org

Blood First Edition Paper, prepublished online November 30, 2006; DOI 10.1182/blood-2006-07-034348

Copyright © 2006 American Society of Hematology

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ABSTRACT

Acute graft-versus-host disease (GVHD) is a major limiting factor in allogeneic hematopoietic

stem cell transplantation (HSCT), and the timing of acute GVHD may affect patient outcomes.

We evaluated the incidence, risk factors, clinical manifestations, and outcomes of hyper-acute

GVHD, defined as that occurring within 14 days after transplant, among 809 consecutive

HSCT at The University of Texas M. D. Anderson Cancer Center. Of 265 patients with grade

II-IV acute GVHD, 27% had biopsy-proven hyper-acute GVHD. Skin involvement was

significantly more common (88% versus 44%) and more severe (stage III-IV, 88% versus 66%)

in the hyper-acute group compared with acute GVHD diagnosed after day 14. On multivariate

analysis, significant risk factors for hyper-acute GVHD included a mismatched related or

matched unrelated donor, a myeloablative conditioning regimen, more than 5 prior

chemotherapy regimens, and donor-recipient sex mismatch. Hyper-acute GVHD was

associated with a significantly lower response rate to first-line therapy and a higher rate of non-

relapse mortality in patients with a mismatched related or matched unrelated donor graft. In

conclusion, hyper-acute GVHD accounts for a substantial proportion of grade II-IV acute

GVHD after HSCT. Patients at high risk or with a diagnosis of hyper-acute GVHD should be

included in clinical studies.

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INTRODUCTION

Acute graft-versus-host disease (GVHD) is one of the major limiting factors in successful

allogeneic hematopoietic stem cell transplantation (HSCT).1,2 Traditionally, acute GVHD has

been defined as a syndrome occurring within the first 100 days following HSCT, with neutrophil

engraftment assumed as a condition for the diagnosis. Acute GVHD defined in this way affects

at least a third of the patients undergoing a matched related (Mrel) donor transplant and a

higher proportion of mismatched and unrelated donor (MUD) transplants.1 As our transplant

practices evolve, it becomes clearer that acute GVHD is better defined as a clinical syndrome

that can occur both early,3 even before engraftment, and late, beyond day 100.4-6 The timing of

acute GVHD may affect the outcome of the disease, with late acute GVHD having a better

outcome than “classic” acute GVHD.6 On the other side of the spectrum, early-onset, or hyper-

acute, GVHD, initially identified in allogeneic transplant recipients without GVHD prophylaxis,7

may be associated with a higher non-relapse mortality rate, particularly in unrelated or

mismatched transplant recipients.3 In this retrospective study, we defined acute GVHD as a

clinical syndrome that can occur any time after an allogeneic infusion, independent of

neutrophil engraftment, and we evaluated the risk factors for and outcomes of hyper-acute

GVHD.

METHODS

Patient eligibility

We identified all consecutive patients who received an allogeneic HSCT as part of prospective

clinical trials at The University of Texas M. D. Anderson Cancer Center between January 1998

and September 2002. The majority of the patients received transplants from human leukocyte

antigen (HLA)–compatible related or unrelated donors serologically matched for HLA-A and -B

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and matched for HLA-DRB1 by high-resolution molecular methods. Patients who received an

umbilical cord transplant or a T-cell-depleted graft were not eligible for this analysis. For

patients who received multiple allogeneic transplants during the study period, only the first

transplant was considered for this analysis, and censoring was performed at the time of the

second transplant.

All patients were treated on clinical protocols, which were reviewed and approved by the M. D.

Anderson Cancer Center Institutional Review Board (IRB). All patients provided written

informed consent before being enrolled in the protocols. IRB approval for this retrospective

chart review was obtained according to institutional guidelines. Demographic and clinical data

were retrieved from the Department of Stem Cell Transplantation and Cellular Therapy

electronic database, which is prospectively updated according to standardized data entry

criteria.

Conditioning regimen and graft-versus-host disease prophylaxis

Conditioning regimens included total body irradiation (TBI)-based myeloablative regimens,

high-dose chemotherapy-based myeloablative regimens, and reduced-intensity regimens.

Preparative regimens were considered myeloablative if they were expected to produce

profound pancytopenia for >28 days without transplantation and if, after transplantation,

hematopoietic recovery was completely donor derived. Reduced-intensity regimens were

defined as those in which hematopoietic recovery was expected to occur within 28 days

without transplantation and, after transplantation, chimerism could be documented in most

patients.8

Almost all patients (98%) received tacrolimus (0.015-0.3 mcg/ Kg/day starting on day –2, with

dose adjustments to maintain blood levels of 5 to 15 ng/dL); methotrexate 5 mg/m2 was given

on days 1, 3, 6, and 11 in bone marrow recipients and on days 1, 3, and 6 in peripheral blood

recipients. Tacrolimus was continued for about 180 days in the absence of disease

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progression or acute GVHD. Pentostatin, steroids, or anti-thymocyte globulin (ATG) were

added to the GVHD prophylaxis regimen as part of studies involving unrelated donor or HLA

mismatched transplants.

Engraftment and chimerism

Engraftment day was defined as the first of three consecutive days in which the patient had an

absolute neutrophil count greater than 0.5 X 109/L. Failure to engraft by day 30 was

considered primary graft failure. Chimerism analysis was performed on days 30 and 100 post-

transplantation and every three months thereafter, according to standard methods.9

Graft-versus-host disease assessment

The diagnosis of acute GVHD was based on clinical signs, positive biopsy results from at least

1 involved organ, and exclusion of other causes of rash, diarrhea, liver function abnormalities,

or other potential manifestations of acute GVHD. The staging and grading of acute GVHD

were performed using the modified Glucksberg consensus criteria.10 Considering a median

time to engraftment of 12 days (range, 7-37 days) in recipients of PBSC grafts and 13 days

(range 5-56) in recipients of bone marrow grafts (overall median 12 days), we defined hyper-

acute GVHD as one occurring within the first 14 days post SCT to encompass time to

engraftment for both PB and BM grafts. The term other acute GVHD was used for cases

developing after day 14, unless otherwise specified.

Treatment of acute graft-versus-host disease

All cases of grade II-IV acute GVHD (except one) were started on methylprednisolone 2

mg/kg/day as per our institutional guidelines. Tacrolimus was continued at blood levels

between 5 and 15 ng/dL. A total of 27 patients received 1 or more additional

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immunosuppressants, including infliximab (n=14), daclizumab (n=6), ATG (n=6), and

basilizumab (n=3).

Evaluation of response to therapy for grade II-IV acute graft-versus-host disease

Responses were assessed for each organ involved. Complete (CR) and partial responses

(PR) were assessed 14 days after the initiation of therapy. A CR was defined as the resolution

of all manifestations of acute GVHD. A PR was a decrease in organ stage by 1. Progressive

disease (PD) was defined as an increase in organ stage by 1 and was evaluated 48 hours

(gastrointestinal [GI] and liver) or 72 hours (skin) after the initiation of corticosteroids. Patients

were considered non-responders (NR) in the absence of CR, PR, or PD 7 days after the

initiation of corticosteroids for skin GVHD or 72 hours after its initiation for GI and liver GVHD.

All responses had a minimal duration of 14 days. Overall response integrated the responses at

all sites (skin, GI, and liver). An overall CR was defined as the resolution of GVHD in all

evaluable organs. PR was any improvement in at least 1 evaluable organ without deterioration

of others; PD was deterioration in at least 1 evaluable organ without improvement of the

others. NR was the absence of any change, or any situation other than CR, PR, or PD.

Statistical methods

Analysis was performed on the basis of outcomes documented by July 2004. Patients who had

a primary graft failure were not eligible for inclusion in the study. Patients who experienced

secondary graft failure after neutrophil engraftment were censored at the time of the secondary

graft failure. Distribution of organ involvement was compared among patients with grade II-IV

hyper-acute and other acute GVHD using the chi-square and Fisher’s exact tests. Prognostic

factors for the occurrence of grade II-IV hyper-acute GVHD and other grade II-IV acute GVHD

and non-relapse mortality among patients diagnosed with grade II-IV acute GVHD were

evaluated using Cox’s regression analysis.11 Observation time was split at the day 14 cut-off

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to evaluate risk factors for hyper-acute and other acute GVHD. Consequently, evaluation of

acute GVHD after day 14 was performed as a landmark analysis starting at day 15 post-HSCT.

Landmark analysis was also used to compare rates of non-relapse mortality after the diagnosis

of grade II-IV acute GVHD. Predictors of response to first-line therapy among patients

diagnosed with grade II-IV acute GVHD were evaluated by logistic regression analysis. The

cumulative incidence of non-relapse mortality was estimated considering death due to

persistence or recurrence of underlying malignancy as competing risk.12 Factors significant at

the 0.1 level on univariate analysis were considered for multivariate analyses using backward

elimination. Two-sided P values of <0.05 were considered significant. Analysis was performed

using STATA 7.0 (Stata Corp., College Station, TX).

RESULTS

Patient characteristics

A total of 809 consecutive patients met the inclusion criteria. Of these, 265 (33%) developed

grade II-IV acute GVHD. Patients’ characteristics are listed in Table 1. The median age was 47

years (range, 19-70 years). Ninety-six percent of patients received an allogeneic stem cell

transplant for hematologic malignancies. Most patients had an HLA-matched sibling donor

transplant (n=488, 60%), and the remainder had HLA-matched unrelated donor (MUD) grafts

(n=253, 31%) and HLA-mismatched related (MMrel) donor grafts (n=68, 8%). The cell source

was peripheral blood stem cells (PBSC) in 53% of patients and bone marrow in 47%. The main

source of stem cells was peripheral blood from Mrel donors (82%) and bone marrow from

MUDs (98%) and MMrel donors (72%). Donor-recipient gender was mismatched (female-to-

male, respectively) in 24% of cases. A third of the patients were in remission of their

malignancy at the time of transplant. The preparative regimen was myeloablative for 477

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patients (59%), and 102 (21%) of these patients received TBI-based conditioning. Three

hundred and thirty-two (41%) patients had reduced-intensity conditioning regimens.

Incidence of acute graft-versus-host disease

Twenty-eight percent (n=73) of the 265 cases of grade II-IV acute GVHD occurred by day 14

after HSCT and are described as hyper-acute GVHD. The majority of hyper-acute GVHD

cases (58/73, 79%) were diagnosed before neutrophil engraftment, and 87% of these cases

(49/56) were diagnosed within 7 days prior to engraftment. Two of the 58 patients who were

diagnosed with acute GVHD before engraftment died without neutrophil recovery. In contrast,

5% (10/192) of cases of acute GVHD diagnosed after day 14 occurred before neutrophil

engraftment. There were a total of 12 cases of grade II-IV acute GVHD which occurred after

disease progression and withdrawal of immunosuppression (3/73 hyper-acute cases and 9/192

other acute GVHD cases). Results of subsequent analyses were unchanged when progression

of the underlying malignancy was considered as a competing risk and observation time was

censored at the time of disease progression before the occurrence of these 12 cases of acute

GVHD (data not shown).

Severity and organ distribution of acute GVHD are summarized on Table 2. Severe, grade III-

IV acute GVHD was seen in 29/73 (40%) patients with hyper-acute GVHD and in 84/192 (44%)

of those with other acute GVHD (P=0.5). The frequency and severity of liver and GI

involvement were comparable in both groups, whereas severe skin involvement was more

common among patients with hyper-acute GVHD (stage III-IV skin GVHD, 71% versus 50%,

P =0.001).

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Risk factors for the development of grade II-IV hyper-acute GVHD

Risk factors for hyper-acute GVHD are summarized in Table 3. On univariate analysis,

significant factors included a MUD (HR=2.2, P=0.002) or MMrel donor (HR=4.1, P <0.001); a

myeloablative conditioning regimen, irrespective of whether it included TBI (HR=3.4, P= 0.002)

or not (HR=3.3, P <0.001); and more than 5 lines of chemotherapy prior to transplantation

(HR=2.1, P =0.04). There was a trend toward higher rates of hyper-acute GVHD in female-to-

male transplants (HR=1.5, P=0.09). Age, gender, disease status, prior autologous transplant,

PBSC (evaluable only among recipients of Mrel donor grafts), and ATG in the conditioning

regimen of MUD transplants (data not shown) did not affect the rate of hyper-acute GVHD.

We evaluated the independent effects of histocompatibility, conditioning regimen, number of

lines of prior chemotherapy, and sex -mismatch in a multivariate model. The results were

consistent with those obtained on univariate analysis, with the exception of female-to-male

transplants, which became statistically significant after adjustment for confounding variables

(HR=1.7, P=0.03).

The impact of T-cell subset cell dose on the incidence of hyper-acute GVHD was only

evaluable in patients who received a PBSC graft from a matched related donor and had

complete cell dose data. A quartile analysis showed a trend towards a lower incidence of

hyper-acute GVHD when either the CD3+ or CD4+ cell doses were within the first quartile

(HRCD3+=0.4, 95% CI 0.1-1.5; HR CD4+=0.3, 95% CI 0.1-1.2). There was no effect for the CD8+

cell dose (HR=0.8, 95% CI 0.3-2.2) or the CD34+ cell dose (HR=1.4, 95% CI 0.6-3.3) in

quartile analysis.

Risk factors for the development of other grade II-IV acute GVHD

Histocompatibility was the only significant predictor of both hyper-acute and other acute

GVHD. Myeloablative regimens showed a trend toward a higher incidence of other acute

GVHD (P =0.05) when TBI- and non-TBI-based regimens were combined. Prior autologous

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transplant had a protective effect for other acute GVHD. However, this effect was only present

in patients who received reduced-intensity conditioning (HR=0.4, P=0.008) but not in those

who received high-dose conditioning (HR=1.1, P=0.8) regimens. On multivariate analysis

adjusting for donor type, prior autologous transplant, conditioning regimen and the interaction

effect between conditioning regimen and prior autologous transplant, only donor type (HR for

MUD =1.6, P=0.004; HR for MMrel donor =1.9, P=0.011) and prior autologous transplant

among recipients of reduced intensity conditioning (HR=0.4, P=0.01) remained significant

predictors for other acute GVHD (Table 3).

Response to therapy in patients with grade II-IV acute GVHD

Response to first-line therapy was assessed among 259 of the 265 patients who developed

grade II-IV GVHD. Six patients diagnosed with other acute GVHD were excluded from the

response evaluation because their responses were not documented or were unknown. Of the

259 patients, 109 (42%) achieved a CR, and 7 (3%) had a PR, resulting in an overall response

rate (CR or PR) of 45%. The overall response rate was significantly lower in the hyper-acute

GVHD group compared with the other acute GVHD group (34% versus 49%, P=0.03). Forty-

seven (64%) of the 73 patients with hyper-acute GVHD and 100 (52%) of the 192 patients with

other acute GVHD required second-line therapy (P= 0.07). This included 4 and 9 patients in

both groups, respectively, who had responded to first-line therapy.

Predictors of response to first-line therapy in patients with grade II-IV acute GVHD

Type of donor and hyper-acute GVHD were the only significant predictors of overall response

to first-line GVHD therapy (Table 4). Because of the strong association between these 2

factors (based on the risk factor analysis described above), we evaluated their independent

effects by grouping patients according to donor histocompatibility and time of onset of acute

GVHD. This analysis showed that patients with a MUD or MMrel donor graft who developed

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hyper-acute GVHD had a significantly lower response rate to first-line therapy (OR=0.3, P=

0.002). Patients who had active disease at transplant showed a trend toward a better response

rate, yet this effect was not significant after adjusting for donor type and hyper-acute GVHD in

multivariate analysis. Patient age, patient gender, donor-recipient sex mismatch, type of

conditioning regimen, and cancer diagnosis had no significant impact on the response rate.

Survival and incidence of chronic GVHD among patients with grade II-IV acute GVHD

Fifty-seven patients (21%) diagnosed with grade II-IV acute GVHD were alive at the time of

this analysis, with a median follow-up of 42 months (range, 5-73 months) since diagnosis of

GVHD. The actuarial survival rate since the diagnosis of grade II-IV GVHD was 21% (95% CI

16%-26%) at the median follow-up time. Overall survival was comparable for patients

diagnosed with hyper-acute (20%, 95% CI 14-27) and other acute GVHD (22%, 95% CI 14-33,

P=0.9). Causes of death are detailed in Table 5. There was no significant difference in non-

relapse mortality for patients with hyper-acute (cumulative incidence 51%) and other acute

GVHD (cumulative incidence 50%, P=0.6). Similarly, there was no significant difference in the

incidence of chronic GVHD between the 2 groups (cumulative incidence, 54% and 59%,

respectively, P=0.2)

Prognostic factors for non-relapse mortality among patients with grade II-IV acute

GVHD

Failure to respond to first-line therapy for treatment of grade II-IV acute GVHD (HR=1.96,

P<0.001) and TBI-based conditioning (HR=1.7, P= 0.02) were the only two statistically

significant predictors on multivariate analysis of non-relapse mortality at 48 months after the

diagnosis of acute GVHD. The cumulative incidence of non-relapse mortality was 60% in non-

responders to first-line therapy versus 39% in responders and 67% in patients who received a

TBI-based conditioning regimen versus 47% in those whose conditioning regimen was not TBI-

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based. Hyper-acute GVHD did not have a significant impact on non-relapse mortality overall

but was associated with a higher mortality rate when it occurred among patients with MMrel or

MUD transplants (cumulative incidence 65%). This rate was significantly higher when

compared with the rate in MMrel or MUD transplants without hyper-acute GVHD or Mrel

transplants (cumulative incidence 47%) (HR=1.7, P 0.01) (Table 6, Figure 1), but it did not

reach statistical significance on multivariate analysis. This increased risk persisted when

evaluated among patients with grade II and III-IV separately, yet it was more pronounced in the

latter group (Table 7). Similarly, the overall non-relapse mortality was higher for grade III-IV

aGVHD (62%) than for grade II (35%) (HR=2.8, P<0.001). Our data showed that patient

gender, donor-patient sex mismatch, patient age, cancer diagnosis, disease status at

transplant, number of prior chemotherapy regimens, cell source, and prior autologous

transplant had no significant impact on the rate of non-relapse mortality.

DISCUSSION

To date, this is the largest study evaluating the incidence of hyper-acute GVHD using standard

criteria for the diagnosis of acute GVHD. The large sample size and the heterogeneity of the

study population allowed us to evaluate risk factors for the occurrence of hyper-acute GVHD,

which is a relatively rare event. Overall, hyper-acute GVHD accounted for 27% of all cases of

grade II-IV acute GVHD diagnosed in our series. A comparison between the incidence of

hyper-acute GVHD in the current study and in the few published reports is not possible

because the definitions vary substantially in terms of both time of onset and clinical

manifestations7,13-16. In our study we used the same diagnostic criteria for hyper-acute GVHD

and acute GVHD occurring post-HSCT.

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Although GVHD is classically related to reactivity of donor T-cells against host tissues, the

severity of acute GVHD is affected by the toxicity of the preparative regimen,4,5 the breakdown

of mucosal barriers, and inflammatory cytokines, in addition to the immune graft-versus-host

reaction. The effects of inflammatory cytokines produced early after the preparative regimen

are likely to contribute to the manifestations of hyper-acute GVHD. Our data support the

hypothesis that preparative regimen toxicity and associated cytokine release would impact

most on the early manifestations of acute GVHD showing that conditioning regimens and

multiple chemotherapy regimens prior to transplant were significant risk factors for hyper-acute

GVHD but not for other acute GVHD. The frequency of hyper-acute GVHD is expected to vary

according to the intensity of the conditioning regimen, thus within and across transplant

centers. On the other hand, HLA disparity was a significant risk factor irrespective of the timing

of GVHD onset. The impact of HLA disparity on the incidence of early-onset acute GVHD has

been reported by Kim et al13 and by Powles and colleagues14,15. Powles et al described the

most severe form of hyper-acute GVHD, which occurred following haploidentical

transplantation and consisted of an abrupt and often fatal clinical syndrome of fever, rash,

massive noncardiogenic pulmonary edema, often with renal failure, and seizures. In the study

of Kim et al,13 an alternative donor source was the only significant predictor of the incidence of

hyper-acute GVHD, which was defined as unexplained fever in addition to skin rash, hepatic

dysfunction, or diarrhea, all occurring before neutrophil engraftment. The authors reported that

conditioning regimen, stem cell source, mononuclear cell dose, or CD34+ cell dose did not

affect the incidence of hyper-acute GVHD among 90 patients, 71 of whom received a graft

from a matched related donor and 19 from an alternative donor. Donor-recipient matching

based on high-resolution HLA typing is likely to lower the incidence of acute GVHD in this

context.

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In the current study, 2 risk factors that have been traditionally associated with acute GVHD-

age (even when evaluated per decade, data not shown) and the infused cell dose of CD-3+, -

4+, -8+, or -34+ with PBSC grafts-did not significantly increase the incidence of GVHD. Our

results show that a prior autologous transplantation had a significant protective effect for the

occurrence of acute GVHD after day 14 but not for hyper-acute GVHD. Most of these patients

received an allogeneic transplant with fludarabine-based reduced-intensity conditioning after

the failure of a prior autologous transplant for the treatment of non-Hodgkin’s lymphoma

(n=35), multiple myeloma (n=18), or Hodgkin’s disease (n=12). The significant protective effect

of a prior autologous transplant was only seen among patients who received reduced-intensity

conditioning and persisted when we restricted the comparison to patients who received a Mrel

or to each one of the diagnoses listed above (data not shown), suggesting that the impact of a

prior autologous transplant among patients who received reduced-intensity conditioning is

independent of donor type or diagnosis. This lower incidence of grade II-IV acute GVHD had

been noted recently by our group in a subset of the non-Hodgkin’s lymphoma17 and multiple

myeloma18 patients included in our current analysis. It is in contrast to prior reports from our

institution19,20 and to reports in the literature of reduced-intensity allogeneic transplant as

salvage therapy after an autologous transplant.21-29 Differences in patient characteristics,

intensity of the conditioning regimen, and the use of donor lymphocyte infusions could account

for this discrepancy. Although the significance of the protective effect is unclear, one could

hypothesize that a prior autologous transplant could induce immunologic changes, leading to

reduced allogeneic reactivity in recipients of a reduced-intensity conditioning regimen.

In addition to having a negative impact on response, hyper-acute GVHD was associated with a

significantly higher rate of non-relapse mortality on univariate analysis in patients who received

MUD or MMrel transplants. This effect could be mainly attributed to resistance to first-line

therapy in this group of patients, which itself is a strong independent predictor of non-relapse

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mortality and would explain the lack of significant effect for hyper-acute GVHD after adjusting

for response to first-line therapy.

The heterogeneity in diagnosis and stage of malignancy of this study population, limits our

ability to assess the graft-versus-malignancy effect. On univariate analysis, patients diagnosed

with grade II-IV hyper-acute GVHD had a lower rate of disease progression compared with

patients with other grade II-IV GVHD, but this trend did not reach statistical significance

(HR=0.6, 95% CI 0.3-1.1). Comprehensive evaluation of the impact of GVHD on disease

progression requires adjustment for confounding variables, which was beyond the scope of

this study.

In conclusion, our data show that acute GVHD comprises a clinical spectrum of GVHD that can

occur early after transplant, even before neutrophil engraftment. The timing of acute GVHD

within this clinical spectrum has a definite impact on the outcomes of patients. Patients at risk

for hyper-acute GVHD should be considered for GVHD prophylaxis within a clinical trial

whenever possible.

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Acknowledgments

The authors would like to acknowledge all research nurses, data managers, nurses, physicians, and physician

assistants for their dedication and hard work in the day-to-day care of our patients.

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Table 1. Patients and transplant characteristics

N 809 (%)

47 (19-70)

543 33 266 67

2 (0-9)

195 24 607 75 7 1

307 38 424 52 31 4 35 4 12 1

539 67 270 33

488 60 253 31 68 8

425 53 384 47

Patient age, years Median (range) >40 ≤40 Prior chemotherapy lines Median (range) Donor/patient gender Female/Male Other Unknown Diagnosis Lymphoid Myeloid Multiple myeloma Solid tumor Other Disease status at transplant Not in remission Remission Donor type Matched related (Mrel) Matched unrelated (MUD) Mismatched related (MMrel) Cell Source PBSC BM Conditioning regimen Reduced intensity 332 41 Fludarabine, Cy/+ATG/+Rituxan/+ Campath 102 13 Fludarabine, Melphalan (140 mg/m2)/ +ATG /+Campath 97 12 Fludarabine, Busulfan/+ATG/+ Campath 50 6 Fludarabine, Idarubicin/Cytarabine 31 4 Cisplatin, Fludarabine,Cytarabine/+ATG 48 6 2CDA,Cytarabine 4 0 High-dose chemotherapy 375 46 Busulfan, Cy/+ATG 143 18 BEAM /+Campath/+Rituxan 59 7 Fludarabine, Busulfan/+ATG 58 7 Fludarabine, Melphalan (180 mg/m2)/+ATG 48 6 F-BAM/ FAM, ATG 32 4 Decitabine/+Busulfan, Cy 15 2 Thiotepa/+Cy,Carmustine/+Melpahalan, Fludarabine/+ Busulfan, Cy 13 2 Cy/+ATG 5 1 Cisplatin, fludarabine, Cytarabine 2 0 High-dose chemotherapy / TBI 102 13 Cy/TBI 52 6 Cy, Thiotepa/TBI 36 4 Fludarabine, Melphalan/TBI 14 2 GVHD prophylaxis Tacrolimus 11 1 Tacrolimus/Methotrexate 759 94 Tacrolimus/Methotrexate/Pentostatin 13 2 Tacrolimus/Methotrexate/Steroids 3 0.4 Tacrolimus -Steroids/-ATG/-ATG & Steroids 3/2/2 1 Cyclosporine/Methotrexate 7 1 Cyclosporine/Steroids 2 0.2 None 7 0.9

PBSC indicates peripheral blood stem cells; BM, bone marrow; Cy, cyclophosphamide; ATG, antithymocyte globulin; CDA, chlorodeoxyadenosine; BEAM, carmustine, etoposide, cytarabine, melphalan; F-BAM, fludarabine, carmustine, cytarabine, melphelan; FAM, flurrouracil, doxorubicin, mitomycin; TBI, total body irradiation; GVHD, graft-versus-host disease.

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Table 2. Grade and stage distribution according to the time of onset of acute GVHD

Hyper-acute Other acute GVHD GVHD P N=73 (%) N=192 (%) Maximun grade II 44 60 108 56 III 11 15 54 28 IV 18 25 30 16 *0.5 Organ distribution Skin stage 0 9 12 66 34 1-2 12 16 29 15 3-4 52 71 97 50 0.001 GI stage 0 37 51 91 47 1-2 19 26 50 26 3-4 17 23 51 27 0.8 Liver stage 0 57 78 155 81 1-2 6 8 17 9 3-4 9 12 20 10 0.9 Unknown 1 1 GVHD indicates graft-versus-host disease *Comparing proportions of grade III-IV acute GVHD

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Table 3. Risk factors for grade II-IV acute GVHD HR 95% CI P HR 95%CI P HR 95% CI P HR 95% CI P Univariate Multivariate Univariate Multivariate

Hyper-acute GVHD Hyper-acute GVHD Other acute GVHD Other acute GVHD

Patient age (years)

>47 0.8 0.5-1.2 0.3 1.1 0.8-1.5 0.5

≤47 1.0 1.0

Patient gender Female 0.9 0.5-1.4 0.5 0.9 0.7-1.2 0.4

Male 1.0 1.0

Donor/patient gender

Female/Male 1.5 0.9-2.5 0.09 1.7 1.1-2.9 0.03 1.1 0.8-1.5 0.7

Other 1.0 1.0 1.0

Unknown

Disease status at transplant

Remission 1.0 1.0

Not in remission 0.9 0.6-1.4 0.7 1.2 0.9-1.7 0.2

Donor type

Matched related 1.0 1.0 1.0 1.0

Matched unrelated 2.2 1.3-3.7 0.002 2.3 1.4-3.9 0.001 1.5 1.13-2.1 0.006 1.6 1.2-2.1 0.004

Mismatched related 4.1 2.1-7.7 <0.001 3.6 1.9-6.9 <0.001 1.9 1.2-3.7 0.01 1.9 1.2-3.2 0.011

Conditioning regimen

Reduced intensity 1.0 1.0

High-dose chemotherapy 3.3 1.8-6.2 <0.001 1.3 0.97-1.8 0.08

TBI 3.4 1.6-7.4 0.002 1.4 0.9-2.3 0.1

HDC/HDC+TBI 3.3 1.8-6.1 <0.001 3.8 2.0-7.0 <0.001 1.3 1.0-1.8 0.05

Diagnosis

Lymphoid 0.9 0.6-1.5 0.8 0.8 0.6-1.1 0.2

Myeloid 1.0 1.0

Multiple myeloma 0.3 0.05-2.5 0.3 1 0.5-2.0 0.9

Solid tumor 2.02 0.9-4.8 0.1 0.7 0.3-1.6 0.4

other 1.8 0.4-7.5 0.4 0.6 0.1-2.3 0.4

Number of prior chemotherapy lines

≤5 1.0 1.0 1.0

>5 2.1 1.04-4.2 0.04 3.1 1.5-6.4 0.002 0.9 0.5-1.8 0.97

Prior autologous transplant

No 1.0 1.0

Yes 0.8 0.3-1.9 0.6 0.5 0.3-0.9 0.03 *0.4 0.2-0.8 0.01

Cell source (in matched related grafts)

BM 1.0 1.0

PBSC 1.3 0.5-3.9 0.6 1.5 0.8-2.5 0.2

GVHD indicates graft-versus-host disease; HR, hazard ratio; TBI, total body irradiation; HDC, high-dose chemotherapy; BM, bone marrow; PBSC, peripheral blood stem cell

* effect among patients receiving reduced-intensity conditioning

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Table 4. Evaluation of predictors of response to first-line therapy for treatment of grade II-IV acute graft-versus-host disease N CR/PR (%) OR 95% CI P OR 95% CI P

259 116 Univariate

Multivariate

Patient age (years) >45 149 73 49 1.5 0.9-2.5 0.1 ≤45 110 43 39 1.0 Patient gender Female 97 39 40 0.7 0.4-1.2 0.25 Male 162 77 47 1.0 Donor/patient gender* Female/Male 73 81 44 1.04 0.6-1.8 0.9 Other 184 33 45 1.0 Acute GVHD Hyper-acute GVHD 73 25 34 0.5 0.3-0.9 0.03 Other acute GVHD 186 91 49 1.0 Donor type Matched related (Mrel) 134 68 51 1.0 Matched unrelated (MUD) 95 41 43 0.7 0.4-1.2 0.2 Mismatched related (MMrel) 30 7 23 0.3 0.1-0.7 0.009 Mismatched related vs other 0.3 0.1-0.8 0.02 Donor type / hyper-acute GVHD Mrel AND no Hyper-acute 106 53 50 1.0 Mrel AND Hyper-acute 28 15 54 1.1 0.5-2.7 0.7 MUD/MMrel AND no Hyper-acute 80 38 47 0.9 0.5-1.6 0.7 MUD/MMrel AND Hyper-acute 45 10 22 0.3 0.1-0.6 0.002 0.3 0.1-0.6 0.002 Cell source (in matched related) PBSC 116 58 50 0.8 0.3-2.2 0.7 BM 18 10 56 1.0 Conditioning regimen Reduced Intensity 83 37 45 1.0 High-Dose Chemotherapy 139 65 47 1.1 0.6-1.9 0.7 TBI 37 14 38 0.8 0.3-1.7 0.5

Diagnosis Myeloid 140 59 42 0.7 0.4-1.3 0.3 Lymphoid 93 46 49 1.0 Multiple Myeloma 10 3 30 0.4 0.1-1.8 0.2 Solid tumor / other 16 8 50 1.02 0.3-2.9 0.96 Disease status at transplant Not in remission 176 86 49 1.7 0.98-2.9 0.06 1.6 0.9-2.7 0.1 Remission 83 30 36 1.0 1.0

Analysis excluding 6 pts with unknown response CR indicates complete remission; PR, partial remission; OR, odds ratio; GVHD, graft-versus-host disease; PBSC, peripheral blood stem cell; BM, bone marrow; TBI, total body irradiation

* Donor gender is unknown for 2 patients

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Table 5. Primary causes of death among patients with grade II-IV acute graft-versus-host disease

Hyper-acute Other acute Overall

GVHD GVHD

N=56 (%) N=152 (%) N=208 (%)

GVHD 27 48 72 47 99 47

Recurrence/persistence of disease 19 34 58 38 77 37

Infection 3 5 11 7 14 7

Organ failure 2 4 2 1 4 2

Other 5 9 9 6 14 7

GVHD indicates graft-versus-host disease

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Table 6. Predictors of non-relapse mortality among patients with grade II-IV acute graft-versus-host disease

N HR 95% CI P HR 95% CI P

265 Univariate Multivariate

Patient age (years)

>40 171 0.9 0.7-1.3 0.7

≤40 94 1.0

Patient gender

Female 101 1.2 0.8-1.7 0.3

Male 164 1.0

Donor/patient gender*

Female/Male 73 1.3 0.96-1.9 0.09

Other 190 1.0

Acute GVHD

Hyper-acute 73 1.1 0.8-1.6 0.6

Other 192 1.0

Donor type

Matched related (Mrel) 137 1.0

Matched unrelated (MUD) 97 1.4 0.98-2.1 0.06

Mismatched related (MMrel) 31 1.7 1.03-2.9 0.04

Donor type / hyper-acute GVHD

Mrel AND no Hyper-acute 109 1.0

Mrel AND Hyper-acute 28 0.6 0.3-1.2 0.1

MUD/MMrel AND No Hyper-acute 83 1.2 0.8-1.8 0.4

MUD/MMrel AND Hyper-acute 45 1.7 1.1-2.7 0.02

MUD/MMrel AND Hyper-acute 45 1.7 1.1-2.6 0.01 1.4 0.9-2.1 0.15

Response to first-line therapy

CR/PR 116 1.0 1.0

Other 143 2.1 1.4-2.9 <0.001 1.96 1.3-2.8 <0.001

Conditioning regimen

Reduced Intensity 88 1.0 1.0

High-Dose Chemotherapy 139 0.8 0.7-1.2 0.4 1.0

TBI 38 1.6 0.98-2.7 0.06 1.7 1.1-2.7 0.02

Number of prior chemotherapy lines*

>5 21 0.8 0.4-1.7 0.6

≤5 243 1.0

Prior autologous transplant

Yes 17 0.6 0.2-1.4 0.2

No 248 1.0

Diagnosis

Lymphoid 95 1.1 0.7-1.5 0.8

Myeloid 143 1.0

Multiple Myeloma 11 0.6 0.2-1.7 0.4

Solid Tumor / Other 16 0.45 0.17-1.2 0.12

Disease status at transplant

Not in remission 181 1.2 0.8-1.7 0.4

Remission 84 1.0

Cell source (among Mrel)

PBSC 118 1.5 0.7-3.2 0.3

BM 19 1.0 HR indicates hazards ratio; GVHD, graft-versus-host disease; CR, complete remission; PR, partial remission; TBI, total body irradiation PBSC, peripheral blood stem cell; BM, bone marrow

* total number of patients differ slightly because of unknown values

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Table 7. Cumulative Incidence of non-relapse mortality at 2 years according to grade of acute GVHD, type of donor and hyper-acute GVHD

Group aGVHD grade II aGVHD grade III-IV

N n (%) % CI NRM HR 95% CI P n (%) %CI NRM HR 95% CI P

Mrel AND no Hyper-acute 109 61 (56%) 31% 1.0 48 (44%) 58% 1.0

Mrel AND Hyper-acute 28 19 (68%) 26% 0.6 0.2-1.6 0.3 9 (32%) 33% 0.8 0.25-2.7 0.8

MUD/MMrel AND No Hyper-acute 82 46 (56%) 35% 1.1 0.6-2.1 0.8 36 (44%) 64% 1.5 0.9-2.6 0.12

MUD/MMrel AND Hyper-acute 45 25 (56%) 54% 1.5 0.8-3.0 0.2 20 (44%) 80% 2.5 1.4-4.6 0.003

*1.6 0.9-2.9 0.1 *2.2 1.2-3.8 0.01

Overall 264 151 (57%) 35% 113 (43%) 62% 2.8 1.9-3.9 <0.001

GVHD indicates graft-versus-host disease; %CI, percent cumulative incidence; NRM, non-relapse mortality; HR, hazard ratio *comparing MUD/MMrel AND Hyper-acute to all other subgroups

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Figure 1. Cumulative incidence of non-relapse mortality according to donor type and the occurrence of grade II-IV hyper-acute graft-

versus-host disease. Hyper-acute graft-versus-host disease was associated with a higher rate of non-relapse mortality when it occurred in patients who

received a mismatched related or a matched unrelated graft (top line) but not in patients who received a matched related graft (lower line). In the absence of

hyper-acute GVHD, the rate of non-relapse mortality was similar for related and unrelated grafts (two middle lines).

0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 75 80 Months since diagnosis of hyper-acute graft-versus-host disease

0.0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1.0

65%

54%

29%

52%

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